To stimulate an interest in science, children need to be exposed to real scientific research to sense the excitement of discovery and gain a feel for what the scientific method truly means. Because of the increasing importance of molecular biology in our daily lives, it is also desirable to expose them to modern molecular biological methods. Finally, in order to be widely disseminated, any program should involve reasonably inexpensive materials. The overall objective of this Phase I proposal is to develop a method to involve school-age children in real scientific research in order to enhance and motivate their educational science curriculum. We have designed the experiments described herein to safely illustrate basic biological ideas in the elementary and secondary school classroom. In brief, the experiments are designed to generate and characterize yeast mutants in a poorly-understood but important large membrane-bound proton-pumping enzyme termed the vacuolar ATPase (V- ATPase). This enzyme is responsible for producing the acidic conditions found in the lumen of such structures as the yeast vacuole, and organelles in higher cells such as the lysosome and the neurosecretory vesicle. Teachers will first be trained over the summer in basic cell biology and in the methods used in the experiments. Teachers will then have their students perform these experiments in the classroom. Students will isolate the (plasmid-borne) gene from each mutant after proving phenotype and send it to the P.I.'s laboratory for sequence determination. All sequences will be shared with the students. Students will be asked to predict the consequences of the mutations on the structure of the enzyme (based on a plausible crystal structure modeled after the known F1FO ATPase structure) and to develop hypotheses about further mutations. At the discretion of the P.I., certain of these hypotheses will then be tested by a subsequent group of students. Because teaching a subject has the effects of solidifying what has been learned, another important aspect of this project will be the involvement of secondary or middle school children to serve as mentors for elementary school children, both in face-to-face learning situations and via the Internet. To share the sequence and assay data, to insure that data collection is occurring in the proper manner, and to provide expert experimental help for both students and teachers, the schoolchildren's classroom will be linked with the laboratory of a scientist (the P.I.) interested and involved in understanding V-ATPase structure and function. We anticipate that dissemination of this program (or similar programs focused on other genes) will be reasonably straightforward. Teachers and students will both gain a strong appreciation of how science really works and we will collect fundamentally interesting mutants in the V-ATPase.